P1328: CHRONIC INFLAMMATION PROMOTES EXTRAMEDULLARY HEMATOPOIESIS VIA UPREGULATION OF MATRIX METHALLOPROTEINASE 9
Authors: Maria Kuzmina, Srdjan Grusanovic, Nataliia Pavliuchenko, Tomas Brdicka, Meritxell Alberich-Jorda
Journal: HemaSphere (2023)

Topic: 23. Hematopoiesis, stem cells and microenvironment Background: Chronic inflammation is a condition characterized by a long-term increased in cytokines and chemokines. It has been reported that chronic inflammatory conditions favor the establishment of new hematopoietic sites outside of the bone marrow (BM), a condition known by the term of extramedullary hematopoiesis (EMH). EMH has been described in pathological conditions in which the BM is inefficient and unable to fulfill the hematopoietic demands. Nevertheless, little is known about how chronic inflammation induces EMH, how it affects hematopoietic stem cells (HSCs), and what are the mechanisms leading to it. Here, we employed a mouse model of Chronic Multifocal Osteomyelitis (CMO) which develops sterile chronic inflammation due to a mutation in the Pstpip2 gene. CMO mice exhibit expansion of the myeloid compartment, swollen paws, and kinks in the tail. Recently, we reported that HSC in the BM of CMO mice present reduced functionality due to hyperactivation of the IL-6/Jak/Stat3 signaling pathway. Aims: The aims of this study are (1) to determine whether CMO mice develop EMH, (2) to functionally characterized the HSCs in EMH sites in CMO mice, and (3) to decipher the mechanisms that promote EMH under chronic inflammatory conditions. Methods: Flow cytometry assays, transplantation assays, generation of chimeras, extreme limiting dilution transplantation assays, in vitro cultivation, RT-PCR, ELISA, gelatinase assays, MMP9 administration in vivo, genome-wide gene expression profile, mass spectrometry. Results: Since EMH is commonly accompanied by the presence of HSCs in circulation, we assessed the presence of Lin-ckit+Sca-1+CD48-CD150+ cells in peripheral blood of CMO mice. We observed increased number of HSCs in circulation of CMO mice, and colony culture assays and murine transplantation experiments demonstrated that these HSCs were functional. Next, we investigated whether CMO mice had HSCs in spleen, a common site of EMH. We observed that CMO mice exhibited higher number of HSCs in spleen, however their functionality was reduced in comparison to wt splenic HSCs. These observations prompted us to search for additional sites of EMH which could further support hematopoiesis under chronic inflammatory conditions. Interestingly, we detected the presence of HSCs in the inflamed paws of CMO mice, and remarkably paw HSCs were able to engraft and reconstitute hematopoiesis in lethally irradiated mice. Further, we investigated which compartment was mediating the occurrence of EMH in CMO mice. Using murine chimeras, we observed that while the BM niche from CMO mice was not mediating the presence of HSCs in the sites of EMH, the CMO immune cells were responsible for the presence of HSCs in peripheral blood, spleen, and paws. Mechanistically, we showed that despite the increased presence of HSC anchoring tools to the BM (CXCR4, VLA-4, and SDF-1), HSCs in CMO conditions were mobilizing to EMH sites. Transcriptomic analysis determined elevated levels of metalloproteinases in CMO mice, and ELISA and gelatinase assays verified the elevated levels and activity of MMP9. Next, we showed that blocking MMP9 activity with SB-3CT prevented mobilization of HSCs in CMO mice. Summary/Conclusion: Altogether, our data indicate that chronic inflammation promotes HSCs mobilization from the BM and leads to the development of EMH. We determined that the immune cells in CMO mice, but not the non-hematopoietic BM niche, mediate HSC mobilization by producing high levels of activated MMP9. The work was partially funded by the National Institute for Cancer Research (Program EXCELES, ID Project No. LX22NPO5102) - Funded by the European Union - Next Generation EU. Keywords: Hematopoietic stem and progenitor cells, Hematopoietic stem cell, Inflammation, Hematopoietic cell transplantation